Nicorandil attenuates cognitive impairment after traumatic brain injury via inhibiting oxidative stress and inflammation: Involvement of BDNF and NGF

Abstract Background and purpose Cognitive impairment is a prevalent adverse consequence of traumatic brain injury (TBI). The neuroprotective effects of nicorandil (N‐(2‐hydroxyethyl)‐nicotinamide nitrate) has been previously documented, yet its protective effects against cognitive dysfunction post‐TBI remain unclear. Hence, the present study was aimed to evaluate whether nicorandil attenuates cognitive dysfunction in TBI rats and the underlying mechanism behind this process. Methods The TBI model was established with a controlled cortical impact (CCI). The effects of nicorandil on cognitive dysfunction of rats with TBI were examined through Novel object recognition (NOR) test, Y‐maze test, and Morris water maze (MWM) task. After behavioral tests, hippocampal tissue was collected for Quantitative real‐time PCR, Western blot analysis, and Enzyme‐linked immunosorbent assay (ELISA) assay. Results We observed that nicorandil administration effectively ameliorates learning and memory impairment in TBI rats. Alongside, nicorandil treatment attenuated oxidative stress in the hippocampus of TBI rats, characterized by the decreased reactive oxygen species generation, malondialdehyde, and protein carbonyls levels, and concurrent promotion of antioxidant‐related factors (including superoxide dismutase, glutathione peroxidase, and catalase) activities. Additionally, nicorandil treatment attenuated the inflammatory response in the hippocampus of TBI rat, as evidenced by the upregulated levels of interleukin (IL)‐1β, IL‐6, and tumor necrosis factor‐α (TNF‐α), as well as the downregulated level of IL‐10. Mechanistically, nicorandil treatment significantly enhanced the mRNA and protein levels of neurotrophic factors, brain‐derived neurotrophic factor (BDNF) and nerve growth factor (NGF) in the hippocampus of TBI rats. Conclusion These findings suggest that nicorandil mitigates cognitive impairment after TBI by suppressing oxidative stress and inflammation, potentially through enhancing BDNF and NGF levels.


INTRODUCTION
Traumatic brain injury (TBI) commonly arises from mechanical forces, such as penetrating trauma or abrupt acceleration/deceleration, resulting in rapid displacement of the brain within the cranial cavity and inflicting acute and severe cerebral damage (Najem et al., 2018).
Emerging evidence demonstrates that TBI is the leading cause of neurologic and long-term neuropsychiatric disability worldwide, closely linked to an increased risk of multiple neurodegenerative diseases including Alzheimer's disease and chronic traumatic encephalopathy (Wilson et al., 2017).Cognitive dysfunction increasingly appares to be a common neurological outcome of TBI (Lai et al., 2022;Paterno et al., 2017).Numerous investigations have consistently demonstrated the upregulation of oxidative stress and inflammatory response in both clinical and preclinical TBI studies, underscoring their significant contributions to subsequent cognitive impairment after TBI (Chio et al., 2022;Corps et al., 2015).Despite the growing recognition of the antioxidant and anti-inflammatory strategies in the management of TBI and/or TBI-associated cognitive impairment is increasingly being recognized (Logsdon et al., 2016), there is no effective pharmaceutical interventions for treating cognitive deficits following TBI until now (Capizzi et al., 2020).Hence, there exists an urgent imperative to develop novel neuroprotective regents that improve cognitive impairment after TBI.
Researches have elucidated that nicorandil promotes cerebral blood flow, attenuates neural death by suppressing oxidative stress and inflammatory effects, and preserves the integrity of the blood-brain barrier, thereby exerting neuroprotective effects (Hosseini et al., 2018;Kotoda et al., 2018).Intriguingly, current investigations suggest that activation of the KATP channel significantly contributes to cerebrovasodilation and plays a pivotal role in autoregulation both of which are impaired in the context of TBI (Pastor et al., 2019).Furthermore, emerging evidence confirms that blockage of KATP channel is closely associated with cognitive impairment (Liu et al., 2021;Moriguchi et al., 2021).These findings raise questions that nicorandil may represent a promising approach for ameliorating cognitive impairment after TBI.
Recent research has elucidated that promotion of neurotrophin levels, specific brain-derived neurotrophic factor (BDNF) (Nguyen et al., 2016) and nerve growth factor (NGF), can attenuate cognitive dysfunction initiated by insults pertinent to the pathophysiology of TBI (Yan et al., 2022;Zhu et al., 2022).Hippocampus, a significant brain region intricately involved in the physiological circuits governing learning and memory, is often injured following TBI (Marzano et al., 2022;Paterno et al., 2017).Thus, the present study aimed to investigate whether nicorandil inhibits cognitive impairment, relying on its antioxidative and anti-inflammatory function.Furthermore, the study delved into the underlying mechanisms, with a particular emphasis on BDNF and NGF levels in the hippocampus of TBI rats.

Controlled cortical impact (CCI) model and drug treatment
A moderate TBI animal model was established using a CCI as previously described protocol with a slight modification (Ma et al., 2019).Briefly, SD rats were anesthetized with sodium pentobarbital (70 mg/kg) and then positioned in a stereotaxic frame.The skull was exposed by mid-longitudinal incision and a 5 mm diameter craniotomy centered at the coronal suture and 3.5 mm lateral to the midline over the left hemisphere was carried out.An electromagnetic impactor device (PinPoint™ Model PCI3000 Precision Cortical Impactor™, Hatteras Instruments, Cary, USA) was attached to a 3.0-mm rounded impacting tip angled at 20-30 • to have a vertical direction to the skull surface.Rats were then exposed to CCI injury with an impact velocity of 3.0 m/s to a depth of 1.0 mm below the dura, and a duration of 180 ms.Sham rats were received a craniotomy except for CCI injury.
Prior to drug administration, rats were randomly divided into four groups (n = 10−12 per group): Sham + Vehicle group, TBI + Vehicle group, TBI + Nicorandil group, and Sham + Nicorandil group.In nicorandil group, rats were orally and daily administrated with nicorandil (7.5 mg/kg) immediately after the surgery until behavioral test, while rats in vehicle group received the same volume of saline.

Novel object recognition (NOR) test
The NOR test procedure was performed on the second day after nicorandil treatment basing on the natural tendency of rats to explore new objects.Briefly, the test was carried out in an open field arena (40 cm × 40 cm × 40 cm).During the 5 min acquisition phase, the rats were allowed to freely move explore the two identical objects (referred to as A and B).Subsequently, in the testing trial, one of the familiar objects (A) was replaced by a novel object (C), which was differed in size from objects A and B, and the rats was allowed to explore for 5 min after a 1 h interval.Object exploration was defined as the animals exhibiting investigative behavior or touching the object with their nose within a 1 cm around the object.The times spent exploring the familiar and novel objects during 5 min period were recorded and analyzed.

Y-maze test
The Y-maze test was employed to assess the short-term spatial recognition memory of the animals, which is based on the rodents' inherent curiosity to explore novel environments.This test was performed on the second day following the NOR test.The Y-maze apparatus composed of three identical arms (40 cm × 20 cm × 10 cm).Each rat was placed in the center of the maze and allowed to freely explore the three arms for a period of 5 min.The sequences of entering arm and the frequency of entries into each arm were meticulously recorded.The alternation behavior (%) was calculated as follows: (number of successful alternations/(total number of arms entries -2) × 100).

Morris water maze (MWM) task
The long-term spatial memory of rodents was assessed using the MWM task, which was conducted one day after the Y-maze test.The water maze consisted of a circular pool with a diameter of 122 cm, featuring a platform with a diameter of 1.5 cm positioned just below the water surface.A fixed camera was positioned above the pool for monitoring purpose.The pool was filled with water (22 ± 1 • C) and divided into four quadrants.The task contains two phases of experimentation.
During the spatial training phase, all rats were tested on four occasions each day for a duration of 5 days.Rats were allowed to freely search for 60 s to find the invisible platform.The time taken to find the platform was recorded as the escape latency.If the rats failed to find the platform within a period of 120 s, they were manually guided to the platform and allowed to remain there for at least 10s.The mean escape latency and the mean swimming speed throughout the five training days were calculated.On the sixth day, the probe trial was performed to evaluate long-term spatial memory capacity of rats.The platform was removed from the pool and the rats were permitted to swim freely for a duration of 60 s.The time spent in the target quadrant and the number of crossings over the target quadrant were recorded.

Detection of reactive oxygen species (ROS) level
The level of ROS in the hippocampal homogenates of rats was detected using the ROS assay kit (Jiancheng Bioengineering Institute, Nan-jing, China) based on the staining of 2,7-dichlorodihydrofluorescein diacetate (DCFH-DA).Briefly, 190 μL of the homogenates and 10 μL of DCFH probe (1 mol/L) were mixed into a 96-well plate at 37 • C for 30 min.The fluorescent intensity (an excitation wavelength of 500 nm, an emission wavelength of 525 nm) was detected using a multidetection microplate reader (BioRad, San Diego, CA, USA).

2.7
Measurement of malondialdehyde (MDA), protein carbonyls and glutathione (GSH) contents, superoxide dismutase (SOD), and catalase (CAT) activities The levels of oxidative stress markers, which included MDA and protein carbonyls, as well as antioxidants markers such as GSH, SOD, and CAT in the hippocampal lysates, were determined using commercially available kits according to the manufacturer's instructions.In brief, 100 mg of hippocampal tissue was cut and thoroughly homogenized in 1 mL of phosphate-buffered saline (PBS) on ice.After centrifugation at 6000 × g for 10 min, the total protein concentration in hippocampal lysates was determined using a BCA reagent kit (CWBio, Beijing, China).The absorbance was measured using a microplate reader (BioRad, San Diego, 535 nm for MDA, 520 nm for protein carbonyls, 412 nm for GSH, 420 nm for SOD, and 405 nm for CAT).The concentration of the respective markers was normalized to the total protein concentration.

Quantification of inflammatory markers levels by enzyme-linked immunosorbent assay (ELISA) assay
Following TBI and nicorandil treatment, the rats were euthanized and the hippocampal tissues were dissected.After homogenization in PBS with a high throughput homogenizer, the homogenates were centrifuged at 12,000 × g for 10 min at 4 • C and the supernatant was collected for further analysis.Then, the levels of interleukin-1 beta (IL-1β), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), and interleukin-10 (IL-10) in the supernatant were quantified following the instructions provided by the ELISA kits.

2.9
Quantitative real-time PCR The total RNA was extracted from hippocampus using the TRIzol

Western blot analysis
The hippocampal tissues (20 mg) were collected, homogenized in pro-

Statistical analysis
Data were analyzed using SPSS 22 (SPSS Inc., Chicago, IL, USA) and expressed as mean ± standard error of the mean (SEM).Statistical significance was determined using a one-way analysis of variance (ANOVA) followed by Tukey's post -hoc test.The escape latency in MWM test was analyzed using two-way repeated-measures ANOVA.
p < .05 was considered significantly different.

Nicorandil attenuates cognitive deficits of TBI rats in the NOR test and Y-maze test
To investigate the impact of nicorandil administration on the learning and memory of TBI rats, the NOR test and Y-maze test were performed on day 1 and day 3 after nicorandil administration, respectively.In the NOR test, compared with the sham rats, the discrimination index of TBI rats was obviously reduced, indicating impaired memory of TBI rats.However, nicorandil treatment significantly increased the discrimination index of TBI rats (Figure 1a).There was no significant difference in the total object exploration time among the four groups (Figure 1b).In the Y-maze test, the correct alternation rates of TBI rats were significantly lower than those of sham rats, whereas the correct alternation rates of rats treated with nicorandil and TBI were higher than those of rats with TBI (Figure 1c).Meanwhile, no observable differences were noted concerning the total number of entries into the arm among the four groups (Figure 1d).These findings suggest that nicorandil treatment ameliorates cognitive impairment induced by TBI.

Nicorandil mitigates spatial learning and memory dysfunction of TBI rats in the Morris water maze test
To further confirm the beneficial effects of nicorandil on cognitive impairment of TBI rats, the Morris water maze (MWM) test was performed to assess spatial learning and memory of rats.A progressive decrease in escape latency was observed across all groups over the course of consecutive training days (Figure 2a).On the fifth day, the TBI rats showed substantial impairment when compared with the sham rats, demonstrated by a prolonged escape latency (Figure 2b) and a reduced percentage of time spent in the target quadrant (Figure 2c) was reduced in TBI rats.However, nicorandil treatment decreased the escape latency (Figure 2b) and increased the percentage of time in the target quadrant in TBI rats (Figure 2c).Moreover, nicorandil treatment upregulated the numbers of crossing target quadrant of TBI rats (Figure 2d).There was no difference in swimming speed among all group during the probe trail test (Figure 2e).Collectively, these findings suggest that nicorandil administration alleviates cognitive impairment of TBI rats.

Nicorandil attenuates oxidative stress with enhanced antioxidants in the hippocampus of TBI rats
Oxidative stress is known to play a crucial role the development of cognitive dysfunction following TBI (Hakiminia et al., 2022).In order to elucidate the underlying mechanism by which nicorandil ameliorates cognitive impairment after TBI, the effects of nicorandil on oxidative injury and antioxidant status during TBI were explored.As shown in Figure 3, the fluorescence intensity of ROS, as well as the levels of lipid peroxidation (MDA) and protein carbonyls in the hippocampus of TBI rats were remarkably increased compared to those in the hippocampus of sham rats (Figure 3a-c).However, the levels of ROS, MDA, and protein carbonyls were significantly reduced in the hippocampus of nicorandil and TBI cotreated rats compared to those in the hippocampus of TBI rats (Figure 3a-c).Furthermore, our findings demonstrated that the activities of GSH, SOD, and CAT, which are the major antioxidant enzymes in maintaining the balance of oxidative stress, were highly reduced in the hippocampus of TBI rats compared to those in sham rats, while nicorandil treatment significantly improved the activities of GSH, SOD, and CAT in the hippocampus of TBI rats (p < .01, Figure 3d and e).Taken together, the results suggested that the beneficial effects of nicorandil in alleviating cognitive dysfunction in TBI rats might be attributed to its ability to mitigate oxidative stress and enhance the antioxidant status in the hippocampus.

Nicorandil inhibits inflammatory response in the hippocampus of TBI rats
It has been reported that inflammatory response contributes to the pathologic and consequent cognitive outcomes after TBI (Bray et al., 2022).To further assess the effect of nicorandil on inflammatory response during TBI, the levels of inflammatory cytokines, including IL-1β, IL-6, TNF-α, and IL-10 were determined by RT-PCR and ELISA.
As shown in Figure 4, the mRNA levels of proinflammatory cytokines, including IL-1β, IL-6, and TNF-α in the hippocampus of the TBI rats were elevated compared to those in the sham rats (Figure 4a-c).However, nicorandil treatment significantly reduced the mRNA levels of IL-1β, IL-6, and TNF-α in the hippocampus of TBI rats (Figure 4a-c).
Significantly, IL-10, a crucial anti-inflammatory cytokine, was obvi-ously reduced in the hippocampus of TBI rats compared to that in the sham rats, but nicorandil treatment remarkably increased the level of hippocampal IL-10 to a certain extent (Figure 4d).There was no significant difference in levels of proinflammatory cytokines and antiinflammatory cytokine in the hippocampus between the nicorandil

Nicorandil promotes hippocampal BDNF and NGF levels in TBI rats
Next, we analyzed the status of the BDNF and NGF in hippocampal tissues across the different experimental groups.RT-PCR and Western blot assays showed that the expression levels of BDNF and NGF mRNA (Figure 5a and b) and proteins (Figure 5c and d) in the hippocampus of TBI rats were lower than those in the sham rats.However, treatment with nicorandil significantly upregulated the expression levels of BDNF and NGF mRNA (Figure 5a and b) and proteins (Figure 5c and d) in the hippocampus of TBI rats.No significant difference was observed between nicorandil treatment and the sham rats.

DISCUSSION
To date, there are no effective available to improving impaired cognition after TBI.Our study explored the effect of nicorandil on cognitive impairment in a rat model of traumatic brain injury (TBI), and to further elucidated the underlying mechanism.First, we observed that nicorandil administration ameliorates learning and memory impair-ment of TBI rats, accompanied by a reduction in oxidative stress and inflammatory response in hippocampus of TBI rats.Mechanistically, these beneficial effects of nicorandil were associated with the upregulation of BDNF and NGF levels in the hippocampus.Collectively, these findings suggested that nicorandil ameliorates cognitive dysfunction following TBI by exerting its antioxidative and anti-inflammatory properties, partially through enhancing hippocampal BDNF and GNF levels.
Cognitive impairment is a prevalent consequence of TBI and contributes to various psychiatric and neurobehavioral deficits associated with TBI (Lai et al., 2022).Multiple mechanisms have been implicated in the development of cognitive dysfunction after TBI, including neuronal apoptosis, oxidative stress, inflammatory responses, and neurotransmitter abnormities (Paterno et al., 2017).Nicorandil, a K + -ATP channel opener, exhibits beneficent effects on the nervous system, including preventing seizures via reducing the excitability of pyramidal neurons (Zhao et al., 2023), decreasing cerebral ischemia/reperfusion (I/R) injury (Owjfard et al., 2020), improving neuronal mitochondrial dysfunction, antioxidative, and anti-apoptosis effects (Ravindran et al., 2017).Notably, nicorandil has also been reported to improve cognitive impairment (Gupta et al., 2016;Singh et al., 2015).Therefore, our study focused on evaluating the effects of nicorandil on cognitive impairment after TBI.Our study revealed that nicorandil treatment effectively ameliorates memory deficits caused by TBI according to the MWM test, which is closely correlated with hippocampal-dependent memory (Brandeis et al., 1989).Similarly, nicorandil treatment also attenuated Considerable evidence supports the involvement of oxidative stress and inflammation in the pathophysiology of memory impairments following TBI (Bing et al., 2023;Chen et al., 2017).Recent studies have demonstrated that persistent oxidative stress and neuroinflammation are commonly observed in the hippocampus of TBI, and inhibiting oxidative stress and neuroinflammation has been associated with the improvement of cognitive dysfunction (Wang et al., 2020;Zhang et al., 2022).For example, the biomarkers of oxidative stress, such as ROS generation and lipid peroxides (MDA) levels, are immediately increased in the hippocampus after TBI, while the elevated scavengers of oxygen radicals, including GSH, SOD, and catalase, significantly reduce oxidative stress and partly reverse the TBI-induced injury and cognitive dysfunction (Li et al., 2019).In addition, previous studies have provided that TBI leads to the release of proinflammatory cytokine and cognitive dysfunction (Chen et al., 2017;Huang et al., 2020).As we know, nicorandil, as a reductor of oxidative damage and/or inflammatory response, has previously been reported to protect and attenuate cognitive dysfunction (Gupta et al., 2016;Singh et al., 2015).Therefore, we further evaluated whether inhibition of oxidative stress and inflammatory response contributes to the protective effects of nicorandil against cognitive impairment following TBI.Our findings revealed an imbalance between oxidation system (upregulation of ROS and MDA levels) and antioxidant system (downregulation of GSH, SOD, and CAT activities) in the hippocampus of TBI rats, which are in accordance with the above reports.However, these effects were blocked by nicorandil administration.Similarly, our results demonstrated that the levels of TNF-α, IL-1β, and IL-6 and/or mRNA, along with a decreased level of the anti-inflammatory cytokine IL-10 in the hippocampus of TBI rats, but these changes of inflammatory cytokines levels were significantly inhibited by nicorandil administration.Taken together, these results suggest that nicorandil exhibits the neuroprotective effects against cognitive impairment following TBI might be related to its antioxidative and anti-neuroinflammation activities.
Neurotrophic factors play a critical role in neural survival, differentiation, function, and plasticity, exerting significant influence on brain development and cognitive function.BDNF and NGF are the members of the neurotrophin family mainly localized in the hippocampus and cortex that supports neural survival and neuronal plasticity asso-ciated with learning and memory function (Chao et al., 2006;Thoenen, 1991).Emerging evidence reveals that the abnormalities in BDNF/NGF synthesis are involved in cognitive dysfunction, while increasing BDNF and NGF levels can restore learning and memory deficiency after brain damage (Ferraguti et al., 2023;Turkmen et al., 2021).Experimental studies have demonstrated that treatment with neurotrophic factors (e.g., NGF, BDNF) can attenuate neuronal death and dysfunction in after brain injury (Giarratana et al., 2019;Zhu et al., 2022).Notably, in our present study, we found that the levels of BDNF and NGF mRNA and proteins were reduced in the hippocampus of TBI rats; for the first time, we demonstrated that nicorandil promotes the levels of BDNF and NGF mRNA and proteins in the hippocampus of TBI rats.These results suggested that upregulation of BDNF and NGF levels may contribute to the protection of nicorandil against cognitive dysfunction after TBI.Previous studies also reveal that BDNF/NGF downregulates cellular oxidative stress and inflammation during brain injury (Oliveira et al., 2022;Scotton et al., 2020), implying the possible involvement of BDNF and NGF in the antioxidative stress and anti-inflammation of nicorandil.
In summary, the present findings provide evidence that nicorandil improves cognitive impairment via inhibiting oxidative stress and neuroinflammation in the hippocampus of TBI rats.Additionally, our study highlights that the beneficent effects of nicorandil are associated with the elevated BDNF and NGF levels.Our findings suggest that treatment with nicorandil could be a strategy to treat TBI-induced neurodegenerative conditions in the brain.
Adult male Sprague Dawley (SD) rats (weighing 200-220 g) were purchased from the medical laboratory animal center of Nanchang University (Nanchang, China) and housed under standard conditions (22-25 • C, 12 h light-dark cycle) with free access to food and water.A period of 7 days was allocated for the rats to acclimate to the laboratory environment prior to the commencement of experiments.All experiments were conducted according to the Guide for the Care and Use of Laboratory Animals (Permit Number: SYPU-IACUC-C2015-0831-203) and approved by the Institutional Animal Care and Use Committees of Nanchang University (No. NCDXSYDWLL-2017619).

F
I G U R E 1 Effects of nicorandil on the behavioral performance of TBI rats in the NOR test and Y-maze test.The NOR test was conducted on day 1 following 30 days of nicorandil treatment.(a) The discrimination index and (b) the total object exploration of rats were calculated (n = 7-9 for each group).The Y-maze test was tested on day 3 after 30 days of nicorandil treatment.(c) The percentage of the correct rate and (d) the total entries number of rats were recorded (n = 7-9 for each group).All data were shown as the mean ± SEM. **p < .01versus the Sham + Vehicle group; # p < and ## p < .01versus the TBI + Vehicle group.

F
Effects of nicorandil on learning and memory of rats in MWM test.(a) Escape latency time during five consecutive days, (b) escape latency time on day 5, (c) time spent in the target quadrant, (d) the number of crossings over the target quadrant, and (e) swimming speed in the MWM test (n = 7-9 for each group) were recorded.All data were shown as the mean ± SEM. **p < .01versus the Sham + Vehicle group; # p < .05 and ## p < .01versus the TBI + Vehicle group.

F
Effects of nicorandil on oxidative stress and antioxidant status in the hippocampus of TBI rats.(a) DCF fluorescence intensity for ROS level, (b) malondialdehyde (MDA) concentration, (c) protein carbonyls level, (d) glutathione (GSH), (e) superoxide dismutase (SOD), and (f) catalase (CAT) activities in the hippocampus were measured (n = 4-6 for each group).All data were shown as the mean ± SEM. *p < .05,**p < .01versus the Sham + Vehicle group; # p < .05 and ## p < .01versus the TBI + Vehicle group.treatment and the sham rats.Collectively, these data demonstrated that nicorandil treatment effectively suppressed the inflammatory response in the hippocampus of TBI rats.

F I G U R E 4
Effects of nicorandil on inflammatory response in the hippocampus of TBI rats.(a) The levels of IL-1β, IL-6, TNF-α, and IL-10 mNRA in the hippocampus were measured by RT-PCR.(b) The levels of IL-1β, (c) IL-6, (d) TNF-α, and (e) IL-10 in the hippocampus were measured by ELISA (n = 4-6 for each group).All data were shown as the mean ± SEM. *p < .05,**p < .01,***p < .001,versus the Sham + Vehicle group; # p < .05 and ## p < .01versus the TBI + Vehicle group.spatial memory and short-term memory impairment of TBI rats, as assessed by the NOR test and the Y-maze test.These findings suggest that nicorandil exerts a protective effect against cognitive impairment following TBI.

F
Effects of nicorandil on BDNF and NGF levels in the hippocampus of TBI rats.The mRNA levels of BDNF (a) and NGF (b) in the hippocampus (n = 4-6 for each group) were detected by RT-PCR (n = 4-6 for each group).(c) The protein expressions of BDNF (d) and NGF (e) in the hippocampus were measured by Western blot analysis (n = 4-6 for each group).All data are shown as the mean ± SEM. *p < .05,**p < .01,versus the Sham + Vehicle group; # p < .05 and ## p < .01versus the TBI + Vehicle group.(TNF-α, IL-1β, and IL-6) in the hippocampus, and administration of antiinflammatory compounds have ability to alleviate TBI-induced injury